The transmission of information from the outside to the inside of a cell or between internal cellular components is essential for cell survival. Small ras-related GTPases are critical components of the cellular signal transduction machinery, disregulation of which can lead to a variety of human diseases. Arl proteins are members of the Arf family of GTPases but their function is not well understood. For example, it is not known whether they act as molecular switches via a nucleotide-binding cycle like other G proteins. Completion of the experiments described in this proposal will help achieve the long-term goal of improving our understanding of the regulation of the Arl GTPases and their function(s) in the cell. The hypothesis that Arl2 acts to regulate multi-protein complexes will be tested by first purifying to homogeneity the Arl2 GTPase activating protein (GAP) activity found in mitochondria. The second aim of this project is to investigate the GAP gene structure and expression profile and to identify related genes in humans and model organisms. The third aim is to examine the function of the protein through biochemical analysis of the GAP activity, subcellular and submitochondrial localization, and identification of its functional domains.